Vaccination has been reported to provide prophylaxis when administered up to four days post-exposure to smallpox, although this effect is not well documented. We have developed a mouse model system to evaluate potential vaccines for prophylaxis after exposure to smallpox. In this model we show that several highly attenuated mutant strains of vaccinia virus can protect when administered intra-nasally one day postexposure to a lethal dose of wt vaccinia virus. In the research proposed in this application we will evaluate numerous strains of vaccinia virus for their ability to protect when administered post-exposure. Route of administration and dose will be optimized. For the most promising strains, mechanism of action will be determined and efficacy will be tested in mice against ectormelia, and in ground squirrels against monkeypox, in preparation for meeting the FDA Animal Efficacy Rule. This work will provide the pre-clinical data for filing an IND fora new post-exposure prophylactic drug for exposure to smallpox.